In previous studies, we have shown that human self-reactive (autoreactive) T cell clones act as helper T cells in unstimulated cultures (cultures lacking polyclonal B cell stimulants such as pokeweed mitogen (PWM)) and as suppressor T cells in stimulated cultures. In the present study, we examine the basis of this dichotomy by measuring cytokine production patterns of the self-reactive T cell clones before and after stimulation. In initial studies, we reiterated previous findings by showing that self-reactive cell clones were functionally distinct from tetanus toxoid (TT) antigen-specific T cell clones. In particular, the self-reactive clones exhibited helper function for B cell Ig production when cultured with B cells alone and suppressor function when cultured with pokeweed mitogen (PWM) or rCD40L plus B cells, whereas TT-specific T cell clones exhibited only helper function. We then showed that the suppressor function was mediated by TGF-beta (not IL-10 or IFN-gamma) in that only the addition of anti- TGF-beta to the cultures reversed the suppression. Parallel studies showed that the self-MHC-reactive T cell clones also inhibit T cell proliferation but in this case, inhibition was mediated by both IL-10 and TGF-beta. In a further series of studies, we investigated the interactions between self-reactive T cell clones and non-T cells which led to suppressor cytokine production. Here we found that pre- stimulation of non-T cells for 8h with PWM results in non-T cells capable of inducing self-reactive T cell clone production of TGF-beta and IL-10; in addition, this pre-stimulation time coincided with peak expression of HLA-DR and costimulatory B7 molecules on the non-T cells. Furthermore, we showed that addition of CTLA-4/Fc to cultures of self- reactive clones plus optimally stimulated non-T cells inhibited induction of TGF-beta production. In summary, these studies show that activated self-reactive T cell clones have the cytokine phenotype of Th3 or Tr1 cells and thus may be important regulatory cells that mediate oral and peripheral tolerance and prevent the occurrence of autoimmunity. - Self-MHC-reactive T cells, Th3 T cells, TGF-beta, IL- 10, CTLA-4, CD80/CD86 - Human Subjects